HIV-1 entry is triggered by interaction of the viral envelope (Env) glycoprotein gp120 with domain 1 (D1) of the T-cell receptor CD4. Binding of CD4 by gp120 induces extensive conformational changes in gp120 leading to formation and exposure of a structure called the co-receptor (coR) binding site, also known as the CD4-induced (CD4i) epitope, in the gp120 protein (Moore et al., Proc Natl Acad Sci USA 100(19): 10598-602, 2003). The bridging sheet of gp120 is a critical component of the coR binding site that is highly conserved across genetically diverse HIV-1 isolates from different clades (Huang et al., Proc Natl Acad Sci USA 101(9):2706-11, 2004). The coR binding site is typically unformed on free (non-CD4-bound) gp120. It forms after attachment of viruses to target cells through CD4 binding. The bridging sheet is highly immunogenic and elicits a class of antibodies known as CD4-induced (CD4i) antibodies in vivo. However, access of full-size Abs to the CD4i epitope (bridging sheet) is sterically restricted during viral entry into cells, most likely because the large size of an Ab cannot access the tight crypt within gp120 where the bridging sheet resides. Thus, most known full-size CD4i Abs do not have potent antiviral activity. Fragments of CD4i Abs that are smaller in size could potentially gain access to the CD4i epitope during viral entry. For example two Ab fragments, known as Fab and scFv, of known CD4i Abs have been shown to significantly inhibit HIV entry more potently than full-size Abs (Labrijn et al., J. Virol. 77: 10557-10565, 2003).
Ibalizumab (iMab) is a potent and broad HIV-1 neutralizing Ab (Jacobson et al., Antimicrob. Agents Chemother. 53:450-457, 2009; Kuritzkes et al., J. Infect. Dis. 189:286-291, 2004). iMab neutralizes HIV by binding mainly to domain 2 (D2) of the CD4 receptor on host T-cells, thus blocking the ability of HIV to use these CD4 receptors to gain entry into T-cells and produce infection (Burkly et al., J. Immunol. 149:1779-178, 1992). In a large panel of primary isolates (more than 100 viruses) tested recently, iMab neutralized 92% of all viruses as defined by 50% inhibition of infection, and 47.4% of viruses as defined by 90% inhibition of infection (FIG. 1). These data, while promising, indicate that there is still a need to further improve the potency and breadth of iMab.
Several antibodies have been reported that target epitopes on HIV Env. m36 polypeptide is a human heavy chain domain Ab fragment that targets the highly conserved, but sterically restricted, CD4i epitope on HIV Env. m36 is a potent and broad cross-reactive HIV-1 inhibitor with CD4i Ab activity in vitro, with a mean IC50 in the hundred nanomolar range (Chen et al., Proc Natl Acad Sci USA 105(44):17121-6, 2008). However, similar to other small-size antibody fragments, the m36 polypeptide is predicted to have a short half life in circulation. A long half life is an important indicator for long-lasting antiviral activity. PG9 is another potent and broad anti-HIV-1 antibody, targeting a quaternary epitope of the HIV envelope trimer (Walker et al., Science 326, 285-289 (2009)). VRC01 is also a potent and broad anti-HIV-1 antibody, targeting a CD4 binding site epitope on the HIV envelope.